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therascreen IDH1/2 RGQ PCR Kit CE

For the detection of 12 IDH1 and IDH2 mutations in glioma
  • Compliance with EU IVD Directive 98/79/EC
  • Accurate detection of 7 IDH1 and 5 IDH2 mutations using PCR clamping
  • Sensitive identification of 3 mutations using ARMS PCR technology
  • A standardized assay with ready-to-use solutions
  • An easy workflow with the Rotor-Gene Q instrument
The therascreen IDH1/2 RGQ PCR Kit enables detection of 12 mutations in the IDH1 and IDH2 genes using real-time PCR on the Rotor-Gene Q MDx 5plex HRM Instrument. The kit provides all necessary reagents, optimized for rapid and sensitive detection of IDH1 and IDH2 mutations.
Cat No./ID: 873011
therascreen IDH1/2 RGQ PCR Kit (20) CE
For 20 reactions: 9 Primer and Probe Mixes, WT Control, Positive Control, Master Mix, Nuclease-Free Water

The therascreen IDH1/2 RGQ PCR Kit CE is intended for in vitro diagnostic use in Europe.


Performance
See the therascreen IDH1/2 RGQ PCR Kit Handbook for detailed performance characteristics.
Principle

The therascreen IDH1/2 RGQ PCR Kit provides reagents to perform 9 separate amplification reactions for detection of 12 mutations:

  • 3 total amplification reactions of codons 132 and 100 of the IDH1 gene and of codon 172 of the IDH2 gene
  • 3 mutation amplification reactions of codons 132 and 100 of the IDH1 gene and of codon 172 of the IDH2 gene
  • 3 mutation-specific amplification reactions of IDH1 R132H, IDH1 R132C, and IDH2 R172K mutations

 

IDH1 and IDH2 mutations detected using the therascreen IDH1/2 RGQ PCR Kit
Gene  Mutation  Base change Cosmic ID

 
 
 IDH1
 Arg132His (R132H  395G>A COSM28746
 Arg132Cys (R132C)  394C>T  COSM28747
 Arg132Ser (R132S)  394C>A  COSM28748
 Arg132Gly (R132G)  394 C>G  COSM28749
 Arg132Leu (R132L)  395G>T  COSM28750
 Arg132Val (R132V)  394_395 CG>GT  COSM28751
 Arg100Gln (R100Q)  299G>A  COSM88208
 
 
 IDH2
 Arg172Lys (R172K)  515G>A  COSM33733
 Arg172Met (R172M)  515G>T  COSM33732
 Arg172Trp (R172W)  514A>T  COSM34039
 Arg172Ser (R172S)  516G>T  COSM34090
 Arg172Gly (R172G)  514A>G COSM33731

Total reaction mixes

The Total Primers and Probe Mixes (PPM-Total) use primers and probes to amplify both mutated and wild-type target sequences.

Mutation detection reaction mixes

The mutation detection primers and probe mixes combine primers and probes, to amplify both mutated and wild-type target sequences, plus an oligonucleotide, 3' blocked with the addition of a phosphate group to prevent elongation (PCR clamping), which is specific to the wild-type target sequence.

When the PCR template contains the wild-type sequence, the 3'-phosphate oligonucleotide will dominate over PCR primer binding due to higher affinity. There is no or low extension by the DNA polymerase and no or low amplification is observed.

When a mutated sequence is present, PCR primer binding will dominate over the 3'-phosphate oligonucleotide binding and amplification will proceed.

Mutation identification reaction mixes

Allele-specific amplification is achieved by ARMS (Amplification Refractory Mutation System), which exploits the ability of the DNA polymerase to distinguish between a match and a mismatch at the 3' end of a PCR primer.

When the PCR primer is fully matched, the amplification proceeds with full efficiency. When the 3' base is mismatched, only low-level background amplification occurs.

Procedure
The therascreen IDH1/2 RGQ PCR Kit provides all necessary reagents for detection of 12 mutations in the IDH1 and IDH2 genes. Nine primer and probe mixes are used to amplify both mutated and wild-type target sequences. Simply add template DNA and the appropriate primer and probe mix to the ready-to-use PCR master mix, and start the reaction on the Rotor-Gene Q MDx 5plex HRM instrument using the optimized cycling program described in the kit handbook. For DNA purification, we recommend using the QIAsymphony DNA Mini Kit on the QIAsymphony SP or the QIAamp DNA FFPE Tissue Kit, following the modified protocols given in the therascreen IDH1/2 RGQ PCR Kit Handbook.
Applications
The therascreen IDH1/2 RGQ PCR Kit is an in vitro diagnostic test based on PCR technology intended for the qualitative detection of 7 mutations in the IDH1 gene and 5 mutations in the IDH2 gene, and for direct identification of 3 major mutations, in DNA extracted from formalin-fixed, paraffin-embedded (FFPE) human brain tissue.
The therascreen IDH1/2 RGQ PCR Kit is intended to be used as an aid for the classification of gliomas.
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